Cell stress response impairs de novo NAD+ biosynthesis in the kidney
Yohan Bignon, Anna Rinaldi, Zahia Nadour, Virginie Poindessous, Ivan Nemazanyy, Olivia Lenoir, Baptiste Fohlen, Pierre Weill-Raynal, Alexandre Hertig, Alexandre Karras, Pierre Galichon, Maarten Naesens, Dany Anglicheau, Pietro E. Cippà, Nicolas Pallet
Yohan Bignon, Anna Rinaldi, Zahia Nadour, Virginie Poindessous, Ivan Nemazanyy, Olivia Lenoir, Baptiste Fohlen, Pierre Weill-Raynal, Alexandre Hertig, Alexandre Karras, Pierre Galichon, Maarten Naesens, Dany Anglicheau, Pietro E. Cippà, Nicolas Pallet
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Research Article Metabolism Nephrology

Cell stress response impairs de novo NAD+ biosynthesis in the kidney

Abstract

The biosynthetic routes leading to de novo nicotinamide adenine dinucleotide (NAD+) production are involved in acute kidney injury (AKI), with a critical role for quinolinate phosphoribosyl transferase (QPRT), a bottleneck enzyme of de novo NAD+ biosynthesis. The molecular mechanisms determining reduced QPRT in AKI, and the role of impaired NAD+ biosynthesis in the progression to chronic kidney disease (CKD), are unknown. We demonstrate that a high urinary quinolinate-to-tryptophan ratio, an indirect indicator of impaired QPRT activity and reduced de novo NAD+ biosynthesis in the kidney, is a clinically applicable early marker of AKI after cardiac surgery and is predictive of progression to CKD in kidney transplant recipients. We also provide evidence that the endoplasmic reticulum (ER) stress response may impair de novo NAD+ biosynthesis by repressing QPRT transcription. In conclusion, NAD+ biosynthesis impairment is an early event in AKI embedded with the ER stress response, and persistent reduction of QPRT expression is associated with AKI to CKD progression. This finding may lead to identification of noninvasive metabolic biomarkers of kidney injury with prognostic and therapeutic implications.

Authors

Yohan Bignon, Anna Rinaldi, Zahia Nadour, Virginie Poindessous, Ivan Nemazanyy, Olivia Lenoir, Baptiste Fohlen, Pierre Weill-Raynal, Alexandre Hertig, Alexandre Karras, Pierre Galichon, Maarten Naesens, Dany Anglicheau, Pietro E. Cippà, Nicolas Pallet

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Figure 5

ER stress reduces QPRT expression in HK2 cells.

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ER stress reduces QPRT expression in HK2 cells. (A and B) Scatter dot pl...
(A and B) Scatter dot plots representing the relative expression of DDIT3 (A) and QPRT (B) transcripts measured by real-time quantitative polymerase chain reaction (RT-qPCR) in HK2 cells incubated 8 hours with 0.25 μM thapsigargin (Tg), with 1 μM dithiothreitol (DTT), with 2.5 μg/mL tunicamycin (Tun), with 5 μg/mL brefeldin A (BFA) or 48 hours in a glucose-deprived culture medium (No Gluc.), and compared with vehicle-treated (DMSO) cells (n = 6–8 replicates). The dashed line represents 1, the reference value of vehicle-treated cells. Bars represent mean ± SEM. (C) Immunoblot representing QPRT, binding immunoglobulin protein (BiP), DDIT3, and tubulin protein expression in HK2 cells 24 hours after incubation with DMSO (vehicle), 1 μM DTT, 2.5 μg/mL Tun, and 5 μg/mL BFA or 48 hours in a glucose-deprived culture medium (No Gluc.). The immunoblot shown is representative of 3 independent experiments. (D and E) Time course analysis of the relative expression of DDIT3 (D) and QPRT (E) transcripts measured by RT-qPCR in HK2 cells incubated with vehicle (DMSO), 0.25 μM Tg, 5 μg/mL BFA, or 2.5 μg/mL Tun for up to 24 hours (4 replicates). Bars represent mean ± SEM. (F) Immunoblot representing the expression of QPRT, BiP, DDIT3, and tubulin proteins in HK2 cells transfected with DDIT3 siRNA (siDDIT3) or with control siRNA and incubated with 2.5 μg/mL Tun, 5 μg/mL BFA, or DMSO (vehicle) for 24 hours. The immunoblot shown is representative of 2 independent experiments. (G) Immunoblot representing the expression of QPRT, BiP, DDIT3, and tubulin proteins in HK2 cells transfected with a pcDNA3.1 vector expressing DDIT3 or an empty vector and incubated with 5 μg/mL BFA or with DMSO (vehicle) for 24 hours. The immunoblot shown is representative of 2 independent experiments.